Soybean variety XB04D04

ABSTRACT

According to the invention, there is provided a novel soybean variety, designated XB04D04. This invention thus relates to the seeds of soybean variety XB04D04, to the plants of soybean XB04D04, to plant parts of soybean variety XB04D04 and to methods for producing a soybean plant produced by crossing soybean variety XB04D04 with another soybean plant, using XB04D04 as either the male or the female parent. This invention also relates to methods for introgressing a transgenic or mutant trait into soybean variety XB04D04 and to the soybean plants and plant parts produced by those methods. This invention also relates to soybean varieties or breeding varieties and plant parts derived from soybean variety XB04D04, to methods for producing other soybean varieties or plant parts derived from soybean variety XB04D04 and to the soybean plants, varieties, and their parts derived from use of those methods. This invention further relates to soybean seeds, plants, and plant parts produced by crossing the soybean variety XB04D04 with another soybean variety.

FIELD OF INVENTION

[0001] This invention is in the field of soybean breeding, specificallyrelating to a soybean variety designated XB04D04.

BACKGROUND OF INVENTION

[0002] The present invention relates to a new and distinctive soybeanvariety, designated XB04D4 which has been the result of years of carefulbreeding and selection as part of a soybean breeding program. There arenumerous steps in the development of any novel, desirable plantgermplasm. Plant breeding begins with the analysis and definition ofproblems and weaknesses of the current germplasm, the establishment ofprogram goals, and the definition of specific breeding objectives. Thenext step is selection of germplasm that possess the traits to meet theprogram goals. The goal is to combine in a single variety an improvedcombination of desirable traits from the parental germplasm. Theseimportant traits may include higher seed yield, resistance to diseasesand insects, tolerance to drought and heat, and better agronomicqualities.

[0003] These, processes, which lead to the final step of marketing anddistribution, can take from six to twelve years from the time the firstcross is made. Therefore, development of new varieties is atime-consuming process that requires precise forward planning, efficientuse of resources, and a minimum of changes in direction.

[0004] Soybean (Glycine max), is an important and valuable field crop.Thus, a continuing goal of soybean breeders is to develop stable, highyielding soybean varieties that are agronomically sound. The reasons forthis goal are to maximize the amount of grain produced on the land usedand to supply food for both animals and humans. To accomplish this goal,the soybean breeder must select and develop soybean plants that have thetraits that result in superior varieties.

[0005] Pioneer soybean research staff creates over 500,000 potential newvarieties each year. Of those new varieties, less than 50 and morecommonly less than 25 are actually selected for commercial use.

[0006] The soybean is the world's leading source of vegetable oil andprotein meal. The oil extracted from soybeans is used for cooking oil,margarine, and salad dressings. Soybean oil is composed of saturated,monounsaturated and polyunsaturated fatty acids. It has a typicalcomposition of 11% palmitic, 4% stearic, 25% oleic, 50% linoleic and 9%linolenic fatty acid content (“Economic Implications of Modified SoybeanTraits Summary Report”, Iowa Soybean Promotion Board & American SoybeanAssociation Special Report 92S, May 1990). Changes in fatty acidcomposition for improved oxidative stability and nutrition areconstantly sought after. Industrial uses of soybean oil which issubjected to further processing include ingredients for paints,plastics, fibers, detergents, cosmetics, and lubricants. Soybean oil maybe split, inter-esterified, sulfurized, epoxidized, polymerized,ethoxylated, or cleaved. Designing and producing soybean oil derivativeswith improved functionality, oliochemistry, is a rapidly growing field.The typical mixture of triglycerides is usually split and separated intopure fatty acids, which are then combined with petroleum-derivedalcohols or acids, nitrogen, sulfonates, chlorine, or with fattyalcohols derived from fats and oils.

[0007] Soybean is also used as a food source for both animals andhumans. Soybean is widely used as a source of protein for animal feedsfor poultry, swine and cattle. During processing of whole soybeans, thefibrous hull is removed and the oil is extracted. The remaining soybeanmeal is a combination of carbohydrates and approximately 50% protein.

[0008] For human consumption soybean meal is made into soybean flourwhich is processed to protein concentrates used for meat extenders orspecialty pet foods. Production of edible protein ingredients fromsoybean offers a healthy, less expensive replacement for animal proteinin meats as well as dairy-type products.

SUMMARY OF INVENTION

[0009] According to the invention, there is provided a novel soybeanvariety, designated XB04D4. This invention thus relates to the seeds ofsoybean variety XB04D4, to the plants of soybean XB04D4, to plant partsof soybean variety XB04D4 and to methods for producing a soybean plantproduced by crossing soybean variety XB04D4 with another soybean plant,using XB04D4 as either the male or the female parent. This inventionalso relates to methods for introgressing a transgenic or mutant traitinto soybean variety XB04D4 and to the soybean plants and plant partsproduced by those methods. This invention also relates to soybeanvarieties or breeding varieties and plant parts derived from soybeanvariety XB04D4, to methods for producing other soybean varieties orplant parts derived from soybean variety XB04D4 and to the soybeanplants, varieties, and their parts derived from use of those methods.This invention further relates to soybean seeds, plants, and plant partsproduced by crossing the soybean variety XB04D4 with another soybeanvariety.

[0010] Definitions

[0011] Certain definitions used in the specification are provided below.Also in the examples which follow, a number of terms are used. In orderto provide a clear and consistent understanding of the specification andclaims, including the scope to be given such terms, the followingdefinitions are provided:

[0012] ALLELE=any of one or more alternative forms of a geneticsequence. In a diploid cell or organism, the two alleles of a givensequence typically occupy corresponding loci on a pair of homologouschromosomes.

[0013] BACKCROSSING=Process in which a breeder crosses a progeny varietyback to one of the parental genotypes one or more times.

[0014] BREEDING=The genetic manipulation of living organisms.

[0015] BREEDING CROSS. A cross to introduce new genetic material into aplant for the development of a new variety. For example, one could crossplant A with plant B, wherein plant B would be genetically differentfrom plant A. After the breeding cross, the resulting F1 plants couldthen be selfed or sibbed for one, two, three or more times (F1, F2, F3,etc.) until a new variety is developed. For clarification, such newvariety would be within a pedigree distance of one breeding cross ofplants A and B. The process described above would be referred to as onebreeding cycle.

[0016] BU/A=Bushels per Acre. The seed yield in bushels/acre is theactual yield of the grain at harvest.

[0017] BSR=Brown Stem Rot Tolerance. This is a visual disease score from1 to 9 comparing all genotypes in a given test. The score is based onleaf symptoms of yellowing and necrosis caused by brown stem rot. Ascore of 9 indicates no symptoms. Visual scores range down to a score of1 which indicates severe symptoms of leaf yellowing and necrosis.

[0018] CW=Canopy Width. This is visual observation of the canopy widthfrom 1 to 9 comparing all genotypes in a given test. The higher thescore the better the canopy width observed.

[0019] CNKR=Stem Canker Tolerance. This is a visual disease score from 1to 9 comparing all genotypes in a given test. The score is based uponpremature plant death. A score of 9 indicates no symptoms, whereas ascore of 1 indicates the entire experimental unit died very early.

[0020] COTYLEDON=A cotyledon is a type of seed leaf. The cotyledoncontains the food storage tissues of the seed.

[0021] ELITE VARIETY=A variety that is sufficiently homozygous andhomogeneous to be used for commercial grain production. An elite varietymay also be used in further breeding.

[0022] EMBRYO=The embryo is the small plant contained within a matureseed.

[0023] EMGSC=Emergence Score. The percentage of emerged plants in a plotrespective to the number of seeds planted.

[0024] F3=This symbol denotes a generation resulting from the selfing ofthe F2 generation along with selection for type and rogueing ofoff-types. The “F” number is a term commonly used in genetics, anddesignates the number of the filial generation. The “F3” generationdenotes the offspring resulting from the selfing or self mating ofmembers of the generation having the next lower “F” number, viz. the F2generation.

[0025] FEC=Iron-deficiency Chlorosis. Plants are scored 1 to 9 based onvisual observations. A score of 1 indicates the plants are dead or dyingfrom iron-deficiency chlorosis, a score of 5 means plants haveintermediate health with some leaf yellowing and a score of 9 means nostunting of the plants or yellowing of the leaves. Plots are usuallyscored in mid July.

[0026] FECL=Iron-deficiency Chlorosis. Plants are scored 1 to 9 based onvisual observations. A score of 1 indicates the plants are dead or dyingfrom iron-deficiency chlorosis, a score of 5 means plants haveintermediate health with some leaf yellowing and a score of 9 means nostunting of the plants or yellowing of the leaves. Plots are scoredaround mid August.

[0027] FEY=Frogeye Tolerance. This is a visual disease score from 1 to 9comparing all genotypes in a given test. The score is based upon leaflesions. A score of 9 indicates no lesions, whereas a score of 1indicates severe leaf necrosis.

[0028] GENOTYPE=Refers to the genetic constitution of a cell ororganism.

[0029] HABIT=This refers to the physical appearance of a plant. It canbe determinate, semi-determinate, intermediate, or indeterminate. Insoybeans, indeterminate varieties are those in which stem growth is notlimited by formation of a reproductive structure (i.e., flowers, podsand seeds) and hence growth continues throughout flowering and duringpart of pod filling. The main stem will develop and set pods over aprolonged period under favorable conditions. In soybeans, determinatevarieties are those in which stem growth ceases at flowering time. Mostflowers develop simultaneously, and most pods fill at approximately thesame time. The terms semi-determinate and intermediate are also used todescribe plant habit and are defined in Bernard, R. L. 1972. “Two genesaffecting stem termination in soybeans.” Crop Science 12:235-239;Woodworth, C. M. 1932. “Genetics and breeding in the improvement of thesoybean.” Bull. Agric. Exp. Stn. (Illinois) 384:297-404; Woodworth, C.M. 1933. “Genetics of the soybean.” J. Am. Soc. Agron. 25:36-51.

[0030] HERBRES=Herbicide Resistance. This indicates that the plant ismore tolerant to the herbicide shown than the level of herbicidetolerance exhibited by wild type plants. A designation of RR indicatestolerance to glyphosate and a designation of STS indicates tolerance tosulfonylurea herbicides.

[0031] HGT=Plant Height. Plant height is taken from the top of the soilto top pod of the plant and is measured in inches.

[0032] HILUM=This refers to the scar left on the seed which marks theplace where the seed was attached to the pod prior to it (the seed)being harvested.

[0033] HYPL=Hypocotyl Elongation. This score indicates the ability ofthe seed to emerge when planted 3″ deep in sand pots and with acontrolled temperature of 25° C. The number of plants that emerge eachday are counted. Based on this data, each genotype is given a 1 to 9score based on its rate of emergence and percent of emergence. A scoreof 9 indicates an excellent rate and percent of emergence, anintermediate score of 5 indicates average ratings and a 1 scoreindicates a very poor rate and percent of emergence.

[0034] HYPOCOTYL=A hypocotyl is the portion of an embryo or seedlingbetween the cotyledons and the root. Therefore, it can be considered atransition zone between shoot and root.

[0035] LDGSEV=Lodging Resistance. Lodging is rated on a scale of 1 to 9.A score of 9 indicates erect plants. A score of 5 indicates plants areleaning at a 45° angle in relation to the ground and a score of 1indicates plants are laying on the ground.

[0036] LEAFLETS=These are part of the plant shoot, and they manufacturefood for the plant by the process of photosynthesis.

[0037] LINKAGE=Refers to a phenomenon wherein alleles on the samechromosome tend to segregate together more often than expected by chanceif their transmission was independent.

[0038] LINKAGE DISEQUILIBRIUM=Refers to a phenomenon wherein allelestend to remain together in linkage groups when segregating from parentsto offspring, with a greater frequency than expected from theirindividual frequencies.

[0039] LLE=Linoleic Acid Percent. Linoleic acid is one of the five mostabundant fatty acids in soybean seeds. It is measured by gaschromatography and is reported as a percent of the total oil content.

[0040] LLN=Linolenic Acid Percent. Linolenic acid is one of the fivemost abundant fatty acids in soybean seeds. It is measured by gaschromatography and is reported as a percent of the total oil content.

[0041] MAT ABS=Absolute Maturity. This term is defined as the length oftime from planting to complete physiological development (maturity). Theperiod from planting until maturity is reached is measured in days,usually in comparison to one or more standard varieties. Plants areconsidered mature when 95% of the pods have reached their mature color.

[0042] MATURITY GROUP=This refers to an agreed-on industry division ofgroups of varieties, based on the zones in which they are adaptedprimarily according to day length or latitude. They consist of very longday length varieties (Groups 000, 00, 0), and extend to very short daylength varieties (Groups VII; VIII, IX, X).

[0043] OIL=Oil Percent. Soybean seeds contain a considerable amount ofoil. Oil is measured by NIR spectrophotometry, and is reported on an asis percentage basis.

[0044] OLC=Oleic Acid Percent. Oleic acid is one of the five mostabundant fatty acids in soybean seeds. It is measured by gaschromatography and is reported as a percent of the total oil content.

[0045] PEDIGREE DISTANCE=Relationship among generations based on theirancestral links as evidenced in pedigrees. May be measured by thedistance of the pedigree from a given starting point in the ancestry.

[0046] PERCENT IDENTITY. Percent identity as used herein refers to thecomparison of the homozygous alleles of two soybean varieties. Percentidentity is determined by comparing a statistically significant numberof the homozygous alleles of two developed varieties. For example, apercent identity of 90% between soybean variety 1 and soybean variety 2means that the two varieties have the same allele at 90% of their loci.

[0047] PERCENT SIMILARITY. Percent similarity as used herein refers tothe comparison of the homozygous alleles of a soybean variety such asXB04D4 with another plant, and if the homozygous allele of XB04D4matches at least one of the alleles from the other plant then they arescored as similar. Percent similarity is determined by comparing astatistically significant number of loci and recording the number ofloci with similar alleles as a percentage. A percent similarity of 90%between XB04D4 and another plant means that XB04D4 matches at least oneof the alleles of the other plant at 90% of the loci.

[0048] PLANT. As used herein, the term “plant” includes reference to animmature or mature whole plant, including a plant from which seed orgrain or anthers have been removed. Seed or embryo that will produce theplant is also considered to be the plant.

[0049] PLANT PARTS. As used herein, the term “plant parts” includesleaves, stems, roots, root tips, anthers, seed, grain, embryo, pollen,ovules, flowers, cotyledon, hypocotyl, pod, flower, shoot and stalk,tissue, cells and the like.

[0050] PLM=Palmitic Acid Percent. Palmitic acid is one of the five mostabundant fatty acids in soybean seeds. It is measured by gaschromatography and is reported as a percent of the total oil content.

[0051] POD=This refers to the fruit of a soybean plant. It consists ofthe hull or shell (pericarp) and the soybean seeds.

[0052] PRT=Phytophthora Tolerance. Tolerance to Phytophthora root rot israted on a scale of 1 to 9, with a score of 9 being the best or highesttolerance ranging down to a score of 1 which indicates the plants haveno tolerance to Phytophthora.

[0053] PRMMAT=Predicted Relative Maturity. Soybean maturities aredivided into relative maturity groups. In the United States the mostcommon maturity groups are 00 through VIII. Within maturity groups 00through V are sub-groups. A sub-group is a tenth of a relative maturitygroup. Within narrow comparisons, the difference of a tenth of arelative maturity group equates very roughly to a day difference inmaturity at harvest.

[0054] PRO=Protein Percent. Soybean seeds contain a considerable amountof protein. Protein is generally measured by NIR spectrophotometry, andis reported on a dry weight basis.

[0055] PUBESCENCE=This refers to a covering of very fine hairs closelyarranged on the leaves, stems and pods of the soybean plant.

[0056] RKI=Root-knot Nematode, Southern. This is a visual disease scorefrom 1 to 9 comparing all genotypes in a given test. The score is basedupon digging plants to visually score the roots for presence or absenceof galling. A score of 9 indicates that there is no galling of theroots, a score of 1 indicates large severe galling cover most of theroot system which results in pre-mature death from decomposing of theroot system.

[0057] RKA=Root-knot Nematode, Peanut. This is a visual disease scorefrom 1 to 9 comparing all genotypes in a given test. The score is basedupon digging plants to look at the roots for presence or absence ofgalling. A score of 9 indicates that there is no galling of the roots, ascore of 1 indicates large severe galling cover most of the root systemwhich results in pre-mature death from decomposing of the root system.

[0058] SCN=Soybean Cyst Nematode Resistance. The score is based onresistance to a particular race of soybean cyst nematode, such as race1, 2, 3, 5 or 14. Scores are visual observations of resistance as versusother genotypes in the test, with a higher score indicating a higherlevel of resistance.

[0059] SD VIG=Seedling Vigor. The score is based on the speed ofemergence of the plants within a plot relative to other plots within anexperiment. A score of 9 indicates that 90% of plants growing haveexpanded first leaves. A score of 1 indicates no plants have expandedfirst leaves.

[0060] SDS=Sudden Death Syndrome. Tolerance to Sudden Death Syndrome israted on a scale of 1 to 9, with a score of 1 being very susceptibleranging up to a score of 9 being tolerant.

[0061] S/LB=Seeds per Pound. Soybean seeds vary in seed size, therefore,the number of seeds required to make up one pound also varies. Thisaffects the pounds of seed required to plant a given area, and can alsoimpact end uses.

[0062] SHATTR=Shattering. This refers to the amount of pod dehiscenceprior to harvest. Pod dehiscence involves seeds falling from the pods tothe soil. This is a visual score from 1 to 9 comparing all genotypeswithin a given test. A score of 9 means pods have not opened and noseeds have fallen out. A score of 5 indicates approximately 50% of thepods have opened, with seeds falling to the ground and a score of 1indicates 100% of the pods are opened.

[0063] SHOOTS=These are a portion of the body of the plant. They consistof stems, petioles and leaves.

[0064] STC=Stearic Acid Percent. Stearic acid is one of the five mostabundant fatty acids in soybean seeds. It is measured by gaschromatography and is reported as a percent of the total oil content.

[0065] WH MD=White Mold Tolerance. This is a visual disease score from 1to 9 comparing all genotypes in a given test. The score is based uponobservations of mycelial growth and death of plants. A score of 9indicates no symptoms. Visual scores of 1 indicate complete death of theexperimental unit.

[0066] Definitions for Area of Adaptability

[0067] When referring to area of adaptability, such term is used todescribe the location with the environmental conditions that would bewell suited for this soybean variety. Area of adaptability is based on anumber of factors, for example: days to maturity, insect resistance,disease resistance, and drought resistance. Area of adaptability doesnot indicate that the soybean variety will grow in every location withinthe area of adaptability or that it will not grow outside the area. Areaof adaptability may also be used to refer to the soil or growingconditions.

[0068] Midwest: Iowa and Missouri

[0069] Heartland: Illinois and the western half of Indiana

[0070] Plains: ⅔ of the eastern parts of South Dakota and Nebraska

[0071] North Central: Minnesota, Wisconsin, the Upper Peninsula ofMichigan, and the eastern half of North Dakota

[0072] Mideast: Michigan, Ohio, and the eastern half of Indiana

[0073] Eastern: Pennsylvania, Delaware, Maryland, Rhode Island, NewJersey, Connecticut, Massachusetts, New York, Vermont, and Maine

[0074] Southern: Virginia, West Virginia, Tennessee, Kentucky, Arkansas,North Carolina, South Carolina, Georgia, Florida, Alabama, Mississippi,and Louisiana

[0075] Western: Texas, Kansas, Colorado, Oklahoma, New Mexico, Arizona,Utah, Nevada, California, Washington, Oregon, Montana, Idaho, Wyoming,the western half of North Dakota, and the western ⅓ South Dakota andNebraska

[0076] PMG infested soils: soils containing Phytophthora sojae

[0077] Narrow rows: 7″ and 15″ row spacing

[0078] High yield environments: areas which lack normal stress forexample they have sufficient rainfall, water drainage, low diseasepressure, and low weed pressure

[0079] Tough environments: areas which have stress challenges, oppositeof a high yield environment

[0080] SCN infected soils: soils containing soybean cyst nematode

[0081] other areas of adaptation include the soybean growing regions ofCanada, tight clay soils, light sandy soils and no-till locations.

DETAILED DESCRIPTION OF INVENTION

[0082] The variety of the invention has shown uniformity and stabilityfor all traits, as described in the following variety descriptioninformation. It has been self-pollinated a sufficient number ofgenerations, with careful attention to uniformity of plant type toensure a sufficient level of homozygosity and phenotypic stability. Thevariety has been increased with continued observation for uniformity. Novariant traits have been observed or are expected.

[0083] Soybean variety XB04D4 is particularly adapted to the NorthCentral and Eastern United States and Canada, and for use in PMGinfected soils.

[0084] Soybean variety XB04D4 demonstrates a valuable combination oftraits, including exceptional yield potential, phytophthora resistanceprovided by the Rps1k gene, and resistance to glyphosate herbicides.

[0085] Soybean variety XB04D4 exhibits a relative maturity of 0 and asubgroup of approximately 4. A variety description of Soybean varietyXB04D4 is provided in Table 1. Traits reported are average values forall locations and years or samples measured.

[0086] Soybean variety XB04D4, being substantially homozygous, can bereproduced by planting seeds of the variety, growing the resultingsoybean plants under self-pollinating or sib-pollinating conditions, andharvesting the resulting seed, using techniques familiar to theagricultural arts. TABLE 1 Variety Description Information XB04D04PERFORMANCE CHARACTERISTICS XB04D04 General Characteristics HerbicideResistance RR, STS RR Avg. Harvest LDGSEV 8 Standability Avg. FieldEmergence EMGSC 7 Avg. Hypocotyl Length HYPLSC 9 Hypocotyl Length L Avg.Canopy Width (9 = wide) CW Avg. Shattering SHATTR Disease/Insect/FungalResistance Phytophthora Race 4 Resistant Phytophthora Race 7 ResistantPhytophthora Race 25 Suscept Avg. Phytophthora PRT 6 Tolerance Avg.Brown Stem Rot BSR Avg. Iron Chlorosis FEC 2 Avg. White Mold WHMD Avg.Cyst Nematode SCN1 Race 1 Avg. Cyst Nematode SCN3 Race 3 Avg. CystNematode SCN5 Race 5 Avg. Cyst Nematode SCN14 Race 14 Avg. Sudden DeathSDS Syndrome Avg. Root-knot RKI Nematode-Southern Avg. Root-knot RKANematode - Peanut Avg. Stem Canker CNKR Avg. Frogeye Leaf Spot FEYOil/Meal Type Avg. Seed Protein (% @ PROT 39.0 Dry Wgt Basis) Avg. SeedOil (% @ Dry OILT 21.5 Wgt Basis) Avg. Seed Size (avg S/LB seeds/lb)Color Characteristics Flower Color FL Purple Pubescence Color PU TawnyHila Color HI Black Pod Color PD Tan Seed Coat Luster SCL Leaf Color LC

[0087] Performance Examples of XB04D04

[0088] In the examples that follow in Table 2, the traits andcharacteristics of soybean variety XB04D04 are given in pairedcomparisons with the Pioneer varieties shown in the following tables.Traits reported are mean values for all locations and years where pairedcomparison data was obtained. TABLE 2A VARIETY COMPARISON DATA FORXB04D04 vs. 90B11 YIELD MATABS LDGSEV FEC EMGSC SDVIG bu/a 60# countscore HGT in score score score OILPCT PROTN G/C count Statistic ABS ABSABS ABS ABS ABS ABS pct ABS pct ABS ABS 90B11 33.1 110.3 6.4 30.3 6.3 86.4 17.9 34.76 14.6 XB04D04 39.8 114.4 8.3 31.3 4.1 8.1 6.7 19.3 33.7616.7 #Locs 19 12 4 4 8 3 3 4 4 3 #Reps 19 12 4 4 8 3 3 4 4 3 #Years 3 33 2 3 2 2 1 1 1 Abs. Diff 6.8 4.1 1.9 1.1 2.2 0.1 0.2 1.39 0.99 2.1 Prob0.000 0.002 0.005 0.443 0.009 0.885 0.184 0.145 0.114 0.066

[0089] TABLE 2B VARIETY COMPARISON DATA FOR XB04D04 vs. 90B11 YIELDMATABS LDGSEV FEC EMGSC SDVIG bu/a 60# count score HGT in score scorescore OILPCT PROTN G/C count Statistic ABS ABS ABS ABS ABS ABS ABS pctABS pct ABS ABS 90B51 40.4 115.7 7.4 32.2 7.2 7.1 5.3 18.71 35.41 16.5XB04D04 42.7 114.4 8.3 31.3 4.1 8.1 6.7 19.3 33.76 16.7 #Locs 20 12 4 48 3 3 4 4 3 #Reps 20 12 4 4 8 3 3 4 4 3 #Years 3 3 3 2 3 2 2 1 1 1 Abs.Diff 2.3 1.3 0.8 0.9 3.1 1 1.3 0.59 1.65 0.2 Prob 0.188 0.161 0.1550.506 0.001 0.423 0.314 0.110 0.013 0.846

[0090] TABLE 2C VARIETY COMPARISON DATA FOR XB04D04 vs. 90B74 YIELDMATABS LDGSEV FEC EMGSC SDVIG bu/a 60# count score HGT in score scorescore OILPCT PROTN G/C count Statistic ABS ABS ABS ABS ABS ABS ABS pctABS pct ABS ABS 90B74 37.4 118.1 8 31.6 7.1 7.2 5.3 19.06 34.14 16.2XB04D04 42.7 114.4 8.3 31.3 4 8.1 6.7 19.3 33.76 16.7 #Locs 20 12 4 4 73 3 4 4 3 #Reps 20 12 4 4 7 3 3 4 4 3 #Years 3 3 3 2 3 2 2 1 1 1 Abs.Diff 5.2 3.7 0.3 0.3 3.2 0.9 1.3 0.24 0.37 0.6 Prob 0.008 0.010 0.6380.919 0.003 0.448 0.195 0.311 0.312 0.416

[0091] TABLE 2D VARIETY COMPARISON DATA FOR XB04D04 vs. 90M20 YIELDMATABS LDGSEV FEC EMGSC SDVIG bu/a 60# count score HGT in score scorescore OILPCT PROTN G/C count Statistic ABS ABS ABS ABS ABS ABS ABS pctABS pct ABS ABS 90M20 38.3 112.2 8 31.3 6.7 7 5.8 19.05 33.91 16 XB04D0442.8 113 8.3 31.3 3.8 8.1 6.7 19.3 33.76 16.7 #Locs 15 9 3 4 7 3 3 4 4 3#Reps 15 9 3 4 7 3 3 4 4 3 #Years 2 2 2 2 2 2 2 1 1 1 Abs. Diff 4.5 0.70.3 0 2.9 1.1 0.9 0.25 0.14 0.7 Prob 0.006 0.442 0.423 1.000 0.001 0.4590.448 0.646 0.787 0.216

Further Embodiments of the Invention

[0092] Genetic Marker Profile Through SSR and First Generation Progeny

[0093] In addition to phenotypic observations, a plant can also beidentified by its genotype. The genotype of a plant can be characterizedthrough a genetic marker profile which can identify plants of the samevariety or a related variety or be used to determine or validate apedigree. Genetic marker profiles can be obtained by techniques such asRestriction Fragment Length Polymorphisms (RFLPs), Randomly AmplifiedPolymorphic DNAs (RAPDs), Arbitrarily Primed Polymerase Chain Reaction(AP-PCR), DNA Amplification Fingerprinting (DAF), Sequence CharacterizedAmplified Regions (SCARs), Amplified Fragment Length Polymorphisms(AFLPs), Simple Sequence Repeats (SSRs) which are also referred to asMicrosatellites, and Single Nucleotide Polymorphisms (SNPs). Forexample, see Cregan et. al, “An Integrated Genetic Linkage Map of theSoybean Genome” Crop Science 39:1464-1490 (1999), and Berry et. al.,Assessing Probability of Ancestry Using Simple Sequence Repeat Profiles:Applications to Maize Inbred Lines and Soybean Varieties” Genetics165:331-342 (2003), each of which are incorporated by reference hereinin their entirety.

[0094] Particular markers used for these purposes are not limited to anyparticular set of markers, but are envisioned to include any type ofmarker and marker profile which provides a means of distinguishingvarieties. One method of comparison is where only the loci for whichXB04D04 is homozygous are used. For example, one set of publiclyavailable markers which could be used to screen and identify varietyXB04D4 is disclosed in Table 3. TABLE 3 Soybean SSR Marker Set MarkersSctt008 Satt372 Satt495 Satt328 Satt582 Satt523 Satt572 Satt389 Satt284Satt165 Satt543 Satt513 Satt042 Satt186 Satt590 Satt300 Sct137 Satt150Satt050 Satt213 Satt567 Satt385 Satt384 Satt540 Satt545 Satt598 Satt175Satt225 Satt204 Satt551 Satt133 Satt602 Satt250 Satt411 Satt452 Satt336Satt233 Satt193 Satt255 Satt327 Satt348 Satt234 Satt421 Satt144 Satt257Satt470 Sat090 Satt358 Satt455 Satt594 Satt259 Satt409 Satt517 Satt420Satt228 Sat117 Satt262 Sct188 Sct187 Satt478 Satt426 Satt353 Satt592Satt509 Satt568 Satt153 Satt251 Sctt009 Satt216 Satt197 Satt279 Satt266Satt303 Satt367 Satt412 Satt577 Satt127 Satt546 Satt467 Sctt012 Satt172Sct034 Satt270 Sat104 Satt304 Satt243 Satt440 Satt601 Satt243 Satt249Satt556 Satt243 Sct046 Satt122 Sct028 Satt596 Satt534 Satt357 Satt380Satt142 Satt532 Satt183 Satt565 Satt221 Satt431 Sct186 Satt383 Satt102Satt451 Satt295 Satt555 Satt227 Satt507 Satt441 Satt432 Satt147 Satt557Satt457 Satt196 Satt475

[0095] Primers and PCR protocols for assaying these and other markersare disclosed in the Soybase (sponsored by the USDA AgriculturalResearch Service and Iowa State University) located at the world wideweb at 129.186.26.94/SSR.html. In addition to being used foridentification of soybean variety XB04D4 and plant parts and plant cellsof variety XB04D4, the genetic profile may be used to identify a soybeanplant produced through the use of XB04D4 or to verify a pedigree forprogeny plants produced through the use of XB04D4. The genetic markerprofile is also useful in breeding and developing backcross conversions.

[0096] The present invention comprises a soybean plant characterized bymolecular and physiological data obtained from the representative sampleof said variety deposited with the ATCC. Further provided by theinvention is a soybean plant formed by the combination of the disclosedsoybean plant or plant cell with another soybean plant or cell andcomprising the homozygous alleles of the variety.

[0097] Means of performing genetic marker profiles using SSRpolymorphisms are well known in the art. SSRs are genetic markers basedon polymorphisms in repeated nucleotide sequences, such asmicrosatellites. A marker system based on SSRs can be highly informativein linkage analysis relative to other marker systems in that multiplealleles may be present. Another advantage of this type of marker isthat, through use of flanking primers, detection of SSRs can beachieved, for example, by the polymerase chain reaction (PCR), therebyeliminating the need for labor-intensive Southern hybridization. The PCRdetection is done by use of two oligonucleotide primers flanking thepolymorphic segment of repetitive DNA. Repeated cycles of heatdenaturation of the DNA followed by annealing of the primers to theircomplementary sequences at low temperatures, and extension of theannealed primers with DNA polymerase, comprise the major part of themethodology.

[0098] Following amplification, markers can be scored by gelelectrophoresis of the amplification products. Scoring of markergenotype is based on the size of the amplified fragment as measured bybase pair weight or molecular weight of the fragment. While variation inthe primer used or in laboratory procedures can affect the reportedweight, relative values should remain constant regardless of thespecific primer or laboratory used. When comparing varieties it ispreferable if all SSR profiles are performed in the same lab.

[0099] Primers used are publicly available and may be found in theSoybase or Cregan supra. See also, PCT Publication No. WO 99/31964Nucleotide Polymorphisms in Soybean, U.S. Pat. No. 6,162,967 PositionalCloning of Soybean Cyst Nematode Resistance Genes, and US 2002/0129402A1Soybean Sudden Death Syndrome Resistant Soybeans and Methods of Breedingand Identifying Resistant Plants, the disclosure of which areincorporated herein by reference.

[0100] The SSR profile of soybean plant XB04D4 can be used to identifyplants comprising XB04D4 as a parent, since such plants will comprisethe same homozygous alleles as XB04D4. Because the soybean variety isessentially homozygous at all relevant loci, most loci should have onlyone type of allele present. In contrast, a genetic marker profile of anF1 progeny should be the sum of those parents, e.g., if one parent washomozygous for allele x at a particular locus, and the other parenthomozygous for allele y at that locus, then the F1 progeny will be xy(heterozygous) at that locus. Subsequent generations of progeny producedby selection and breeding are expected to be of genotype x (homozygous),y (homozygous), or xy (heterozygous) for that locus position. When theF1 plant is selfed or sibbed for successive filial generations, thelocus should be either x or y for that position.

[0101] In addition, plants and plant parts substantially benefiting fromthe use of XB04D4 in their development, such as XB04D4 comprising abackcross conversion, transgene, or genetic sterility factor, may beidentified by having a molecular marker profile with a high percentidentity to XB04D4. Such a percent identity might be 95%, 96%, 97%, 98%,99%, 99.5% or 99.9% identical to XB04D4.

[0102] The SSR profile of XB04D4 also can be used to identifyessentially derived varieties and other progeny varieties developed fromthe use of XB04D4, as well as cells and other plant parts thereof. Suchplants may be developed using the markers identified in WO 00/31964,U.S. Pat. No. 6,162,967 and US2002/0129402A1. Progeny plants and plantparts produced using XB04D4 may be identified by having a molecularmarker profile of at least 25%, 30%, 35%, 40%, 45%, 50%, 55%, 60%, 65%,70%, 75%, 79%, 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%,91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or 99.5% geneticcontribution from soybean variety, as measured by either percentidentity or percent similarity. Such progeny may be furthercharacterized as being within a pedigree distance of XB04D4, such aswithin 1, 2, 3, 4 or 5 or less cross-pollinations to a soybean plantother than XB04D4 or a plant that has XB04D4 as a progenitor. Uniquemolecular profiles may be identified with other molecular tools such asSNPs and RFLPs.

[0103] While determining the SSR genetic marker profile of the plantsdescribed supra, several unique SSR profiles may also be identifiedwhich did not appear in either parent of such plant. Such unique SSRprofiles may arise during the breeding process from recombination ormutation. A combination of several unique alleles provides a means ofidentifying a plant variety, an F1 progeny produced from such variety,and progeny produced from such variety.

[0104] Introduction of a New Trait or Locus into XB04D4

[0105] Variety XB04D4 represents a new base genetic variety into which anew locus or trait may be introgressed. Direct transformation andbackcrossing represent two important methods that can be used toaccomplish such an introgression. The term backcross conversion andsingle locus conversion are used interchangeably to designate theproduct of a backcrossing program.

[0106] Backcross Conversions of XB04D4

[0107] A backcross conversion of XB04D4 occurs when DNA sequences areintroduced through backcrossing (Hallauer et al, 1988), with XB04D4utilized as the recurrent parent. Both naturally occurring andtransgenic DNA sequences may be introduced through backcrossingtechniques. A backcross conversion may produce a plant with a trait orlocus conversion in at least two or more backcrosses, including at least2 crosses, at least 3 crosses, at least 4 crosses, at least 5 crossesand the like. Molecular marker assisted breeding or selection may beutilized to reduce the number of backcrosses necessary to achieve thebackcross conversion. For example, see Openshaw, S. J. et al.,Marker-assisted Selection in Backcross Breeding. In: ProceedingsSymposium of the Analysis of Molecular Data, August 1994, Crop ScienceSociety of America, Corvallis, Oreg., where it is demonstrated that abackcross conversion can be made in as few as two backcrosses.

[0108] The complexity of the backcross conversion method depends on thetype of trait being transferred (single genes or closely linked genes asvs. unlinked genes), the level of expression of the trait, the type ofinheritance (cytoplasmic or nuclear) and the types of parents includedin the cross. It is understood by those of ordinary skill in the artthat for single gene traits that are relatively easy to classify, thebackcross method is effective and relatively easy to manage. (SeeHallauer et al. in Corn and Corn Improvement, Sprague and Dudley, ThirdEd. 1998). Desired traits that may be transferred through backcrossconversion include, but are not limited to, sterility (nuclear andcytoplasmic), fertility restoration, nutritional enhancements, droughttolerance, nitrogen utilization, altered fatty acid profile, lowphytate, industrial enhancements, disease resistance (bacterial, fungalor viral), insect resistance and herbicide resistance. In addition, anintrogression site itself, such as an FRT site, Lox site or other sitespecific integration site, may be inserted by backcrossing and utilizedfor direct insertion of one or more genes of interest into a specificplant variety. In some embodiments of the invention, the number of locithat may be backcrossed into XB04D4 is at least 1, 2, 3, 4, or 5 and/orno more than 6, 5, 4, 3, or 2. A single locus may contain severaltransgenes, such as a transgene for disease resistance that, in the sameexpression vector, also contains a transgene for herbicide resistance.The gene for herbicide resistance may be used as a selectable markerand/or as a phenotypic trait. A single locus conversion of site specificintegration system allows for the integration of multiple genes at theconverted loci.

[0109] The backcross conversion may result from either the transfer of adominant allele or a recessive allele. Selection of progeny containingthe trait of interest is accomplished by direct selection for a traitassociated with a dominant allele. Transgenes transferred viabackcrossing typically function as a dominant single gene trait and arerelatively easy to classify. Selection of progeny for a trait that istransferred via a recessive allele requires growing and selfing thefirst backcross generation to determine which plants carry the recessivealleles. Recessive traits may require additional progeny testing insuccessive backcross generations to determine the presence of the locusof interest. The last backcross generation is usually selfed to givepure breeding progeny for the gene(s) being transferred, although abackcross conversion with a stably introgressed trait may also bemaintained by further backcrossing to the recurrent parent withselection for the converted trait.

[0110] Along with selection for the trait of interest, progeny areselected for the phenotype of the recurrent parent. The backcross is aform of inbreeding, and the features of the recurrent parent areautomatically recovered after successive backcrosses. Poehlman, BreedingField Crops, P. 204 (1987). Poehlman suggests from one to four or morebackcrosses, but as noted above, the number of backcrosses necessary canbe reduced with the use of molecular markers. Other factors, such as agenetically similar donor parent, may also reduce the number ofbackcrosses necessary. As noted by Poehlman, backcrossing is easiest forsimply inherited, dominant and easily recognized traits.

[0111] One process for adding or modifying a trait or locus in soybeanvariety XB04D4 comprises crossing XB04D4 plants grown from XB04D4 seedwith plants of another soybean variety that comprise the desired traitor locus, selecting F1 progeny plants that comprise the desired trait orlocus to produce selected F1 progeny plants, crossing the selectedprogeny plants with the XB04D4 plants to produce backcross progenyplants, selecting for backcross progeny plants that have the desiredtrait or locus and the morphological characteristics of soybean varietyXB04D4 to produce selected backcross progeny plants; and backcrossing toXB04D4 three or more times in succession to produce selected fourth orhigher backcross progeny plants that comprise said trait or locus. Themodified XB04D4 may be further characterized as having the physiologicaland morphological characteristics of soybean variety XB04D4 listed inTable 1 as determined at the 5% significance level when grown in thesame environmental conditions and/or may be characterized by percentsimilarity or identity to XB04D04 as determined by SSR markers. Theabove method may be utilized with fewer backcrosses in appropriatesituations, such as when the donor parent is highly related or markersare used in the selection step. Desired traits that may be used includethose nucleic acids known in the art, some of which are listed herein,that will affect traits through nucleic acid expression or inhibition.Desired loci include the introgression of FRT, Lox and other sites forsite specific integration, which may also affect a desired trait if afunctional nucleic acid is inserted at the integration site.

[0112] In addition, the above process and other similar processesdescribed herein may be used to produce first generation progeny soybeanseed by adding a step at the end of the process that comprises crossingXB04D4 with the introgressed trait or locus with a different soybeanplant and harvesting the resultant first generation progeny soybeanseed.

[0113] Transformation

[0114] The advent of new molecular biological techniques has allowed theisolation and characterization of genetic elements with specificfunctions, such as encoding specific protein products. Scientists in thefield of plant biology developed a strong interest in engineering thegenome of plants to contain and express foreign genetic elements, oradditional, or modified versions of native or endogenous geneticelements in order to alter the traits of a plant in a specific manner.Any DNA sequences, whether from a different species or from the samespecies, that are inserted into the genome using transformation arereferred to herein collectively as “transgenes”. In some embodiments ofthe invention, a transformed variant of XB04D4 may contain at least onetransgene but could contain at least 1, 2, 3, 4, 5, 6, 7, 8, 9, 10and/or no more than 15, 14, 13, 12, 11, 10, 9, 8, 7, 6, 5, 4, 3, or 2.Over the last fifteen to twenty years several methods for producingtransgenic plants have been developed, and the present invention alsorelates to transformed versions of the claimed soybean variety XB04D4.

[0115] One embodiment of the invention is a process for producingsoybean variety XB04D04 further comprising a desired trait, said processcomprising transforming a soybean plant of variety XB04D04 with atransgene that confers a desired trait. Another embodiment is theproduct produced by this process. In one embodiment the desired traitmay be one or more of herbicide resistance, insect resistance, diseaseresistance, decreased phytate, or modified fatty acid or carbohydratemetabolism. The specific gene may be any known in the art or listedherein, including; a polynucleotide conferring resistance toimidazolinone, sulfonylurea, glyphosate, glufosinate, triazine andbenzonitrile; a polynucleotide encoding a bacillus thuringiensispolypeptide, a polynucleotide encoding phytase, FAD-2, FAD-3, galactinolsynthase or a raffinose synthetic enzyme; or a polynucleotide conferringresistance to soybean cyst nematode, brown stem rot, phytophthora rootrot, soybean mosaic virus or sudden death syndrome.

[0116] Numerous methods for plant transformation have been developed,including biological and physical plant transformation protocols. See,for example, Miki et al., “Procedures for Introducing Foreign DNA intoPlants” in Methods in Plant Molecular Biology and Biotechnology, Glick,B. R. and Thompson, J. E. Eds. (CRC Press, Inc., Boca Raton, 1993) pages67-88 and Armstrong, “The First Decade of Maize Transformation: A Reviewand Future Perspective” (Maydica 44:101-109, 1999). In addition,expression vectors and in vitro culture methods for plant cell or tissuetransformation and regeneration of plants are available. See, forexample, Gruber et al., “Vectors for Plant Transformation” in Methods inPlant Molecular Biology and Biotechnology, Glick, B. R. and Thompson, J.E. Eds. (CRC Press, Inc., Boca Raton, 1993) pages 89-119.

[0117] The most prevalent types of plant transformation involve theconstruction of an expression vector. Such a vector comprises a DNAsequence that contains a gene under the control of or operatively linkedto a regulatory element, for example a promoter. The vector may containone or more genes and one or more regulatory elements.

[0118] A genetic trait which has been engineered into the genome of aparticular soybean plant using transformation techniques, could be movedinto the genome of another variety using traditional breeding techniquesthat are well known in the plant breeding arts. For example, abackcrossing approach may be used to move a transgene from a transformedsoybean variety into an already developed soybean variety, and theresulting backcross conversion plant would then comprise thetransgene(s).

[0119] Various genetic elements can be introduced into the plant genomeusing transformation. These elements include, but are not limited togenes; coding sequences; inducible, constitutive, and tissue specificpromoters; enhancing sequences; and signal and targeting sequences. Forexample, see the traits, genes and transformation methods listed in U.S.Pat. No. 6,118,055.

[0120] With transgenic plants according to the present invention, aforeign protein can be produced in commercial quantities. Thus,techniques for the selection and propagation of transformed plants,which are well understood in the art, yield a plurality of transgenicplants that are harvested in a conventional manner, and a foreignprotein then can be extracted from a tissue of interest or from totalbiomass. Protein extraction from plant biomass can be accomplished byknown methods which are discussed, for example, by Heney and Orr, Anal.Biochem. 114: 92-6 (1981).

[0121] A genetic map can be generated, primarily via conventionalRestriction Fragment Length Polymorphisms (RFLP), Polymerase ChainReaction (PCR) analysis, Simple Sequence Repeats (SSR) and SingleNucleotide Polymorphisms (SNP) that identifies the approximatechromosomal location of the integrated DNA molecule. For exemplarymethodologies in this regard, see Glick and Thompson, METHODS IN PLANTMOLECULAR BIOLOGY AND BIOTECHNOLOGY 269-284 (CRC Press, Boca Raton,1993).

[0122] Wang et al. discuss “Large Scale Identification, Mapping andGenotyping of Single-Nucleotide Polymorphisms in the Human Genome”,Science, 280:1077-1082, 1998, and similar capabilities are becomingincreasingly available for the soybean genome. Map informationconcerning chromosomal location is useful for proprietary protection ofa subject transgenic plant. If unauthorized propagation is undertakenand crosses made with other germplasm, the map of the integration regioncan be compared to similar maps for suspect plants to determine if thelatter have a common parentage with the subject plant. Map comparisonswould involve hybridizations, RFLP, PCR, SSR and sequencing, all ofwhich are conventional techniques. SNPs may also be used alone or incombination with other techniques.

[0123] Likewise, by means of the present invention, plants can begenetically engineered to express various phenotypes of agronomicinterest. Through the transformation of soybean the expression of genescan be modulated to enhance disease resistance, insect resistance,herbicide resistance, agronomic, grain quality and other traits.Transformation can also be used to insert DNA sequences which control orhelp control male-sterility. DNA sequences native to soybean as well asnon-native DNA sequences can be transformed into soybean and used tomodulate levels of native or non-native proteins. Various promoters,targeting sequences, enhancing sequences, and other DNA sequences can beinserted into the genome for the purpose of modulating the expression ofproteins. Reduction of the activity of specific genes (also known asgene silencing, or gene suppression) is desirable for several aspects ofgenetic engineering in plants.

[0124] Many techniques for gene silencing are well known to one of skillin the art, including but not limited to antisense technology (see,e.g., Sheehy et al. (1988) PNAS USA 85:8805-8809; and U.S. Pat. Nos.5,107,065; 5,453,566; and 5,759,829); co-suppression (e.g., Taylor(1997) Plant Cell 9:1245; Jorgensen (1990) Trends Biotech. 8(12):340-344; Flavell (1994) PNAS USA 91:3490-3496; Finnegan et al. (1994)Bio/Technology 12: 883-888; and Neuhuber et al. (1994) Mol. Gen. Genet.244:230-241); RNA interference (Napoli et al. (1990) Plant Cell2:279-289; U.S. Pat. No. 5,034,323; Sharp (1999) Genes Dev. 13:139-141;Zamore et al. (2000) Cell 101:25-33; and Montgomery et al. (1998) PNASUSA 95:15502-15507), virus-induced gene silencing (Burton, et al. (2000)Plant Cell 12:691-705; and Baulcombe (1999) Curr. Op. Plant Bio.2:109-113); target-RNA-specific ribozymes (Haseloff et al. (1988) Nature334: 585-591); hairpin structures (Smith et al. (2000) Nature407:319-320; WO 99/53050; and WO 98/53083); ribozymes (Steinecke et al.((1992) EMBO J. 11:1525; and Perriman et al. ((1993) Antisense Res. Dev.3:253); oligonucleotide mediated targeted modification (e.g., WO03/076574 and WO 99/25853); Zn-finger targeted molecules (e.g., WO01/52620; WO 03/048345; and WO 00/42219); and other methods orcombinations of the above methods known to those of skill in the art.

[0125] Exemplary transgenes useful for genetic engineering include, butare not limited to, those categorized below.

[0126] 1. Transgenes that Confer Resistance to Insects or Disease andthat Encode:

[0127] (A) Plant disease resistance genes. Plant defenses are oftenactivated by specific interaction between the product of a diseaseresistance gene (R) in the plant and the product of a correspondingavirulence (Avr) gene in the pathogen. A plant variety can betransformed with cloned resistance gene to engineer plants that areresistant to specific pathogen strains. See, for example Jones et al.,Science 266: 789 (1994) (cloning of the tomato Cf-9 gene for resistanceto Cladosporium fulvum); Martin et al., Science 262: 1432 (1993) (tomatoPto gene for resistance to Pseudomonas syringae pv. tomato encodes aprotein kinase); Mindrinos et al., Cell 78: 1089 (1994) (ArabidopsisRSP2 gene for resistance to Pseudomonas syringae). A plant resistant toa disease is one that is more resistant to a pathogen as compared to thewild type plant.

[0128] (B) A Bacillus thuringiensis protein, a derivative thereof or asynthetic polypeptide modeled thereon. See, for example, Geiser et al.,Gene 48: 109 (1986), who disclose the cloning and nucleotide sequence ofa Bt delta-endotoxin gene. Moreover, DNA molecules encodingdelta-endotoxin genes can be purchased from American Type CultureCollection (Rockville, Md.), for example, under ATCC Accession Nos.40098, 67136, 31995 and 31998. Other examples of Bacillus thuringiensistransgenes being genetically engineered are given in the followingpatents and patent applications and hereby are incorporated by referencefor this purpose: U.S. Pat. Nos. 5,188,960; 5,689,052; 5,880,275; WO91/114778; WO 99/31248; WO 01/12731; WO 99/24581; WO 97/40162 and U.S.application Ser. Nos. 10/032,717; 10/414,637; and 10/606,320.

[0129] (C) An insect-specific hormone or pheromone such as anecdysteroid and juvenile hormone, a variant thereof, a mimetic basedthereon, or an antagonist or agonist thereof. See, for example, thedisclosure by Hammock et al., Nature 344: 458 (1990), of baculovirusexpression of cloned juvenile hormone esterase, an inactivator ofjuvenile hormone.

[0130] (D) An insect-specific peptide which, upon expression, disruptsthe physiology of the affected pest. For example, see the disclosures ofRegan, J. Biol. Chem. 269: 9 (1994) (expression cloning yields DNAcoding for insect diuretic hormone receptor), and Pratt et al., Biochem.Biophys. Res. Comm. 163: 1243 (1989) (an allostatin is identified inDiploptera puntata). See also U.S. Pat. No. 5,266,317 to Tomalski etal., who disclose genes encoding insect-specific toxins.

[0131] (E) An enzyme responsible for an hyperaccumulation of amonterpene, a sesquiterpene, a steroid, hydroxamic acid, aphenylpropanoid derivative or another non-protein molecule withinsecticidal activity.

[0132] (F) An enzyme involved in the modification, including thepost-translational modification, of a biologically active molecule; forexample, a glycolytic enzyme, a proteolytic enzyme, a lipolytic enzyme,a nuclease, a cyclase, a transaminase, an esterase, a hydrolase, aphosphatase, a kinase, a phosphorylase, a polymerase, an elastase, achitinase and a glucanase, whether natural or synthetic. See PCTapplication WO 93/02197 in the name of Scott et al., which discloses thenucleotide sequence of a callase gene. DNA molecules which containchitinase-encoding sequences can be obtained, for example, from the ATCCunder Accession Nos. 39637 and 67152. See also Kramer et al., InsectBiochem. Molec. Biol. 23: 691 (1993), who teach the nucleotide sequenceof a cDNA encoding tobacco hookworm chitinase, and Kawalleck et al.,Plant Molec. Biol. 21: 673 (1993), who provide the nucleotide sequenceof the parsley ubi4-2 polyubiquitin gene.

[0133] (G) A molecule that stimulates signal transduction. For example,see the disclosure by Botella et al., Plant Molec. Biol. 24: 757 (1994),of nucleotide sequences for mung bean calmodulin cDNA clones, and Griesset al., Plant Physiol. 104: 1467 (1994), who provide the nucleotidesequence of a maize calmodulin cDNA clone.

[0134] (H) A hydrophobic moment peptide. See PCT application WO 95/16776(disclosure of peptide derivatives of Tachyplesin which inhibit fungalplant pathogens) and PCT application WO 95/18855 (teaches syntheticantimicrobial peptides that confer disease resistance), the respectivecontents of which are hereby incorporated by reference for this purpose.

[0135] (I) A membrane permease, a channel former or a channel blocker.For example, see the disclosure by Jaynes et al., Plant Sci. 89: 43(1993), of heterologous expression of a cecropin-beta lytic peptideanalog to render transgenic tobacco plants resistant to Pseudomonassolanacearum.

[0136] (J) A viral-invasive protein or a complex toxin derivedtherefrom. For example, the accumulation of viral coat proteins intransformed plant cells imparts resistance to viral infection and/ordisease development effected by the virus from which the coat proteingene is derived, as well as by related viruses. See Beachy et al., Ann.Rev. Phytopathol. 28: 451 (1990). Coat protein-mediated resistance hasbeen conferred upon transformed plants against alfalfa mosaic virus,cucumber mosaic virus, tobacco streak virus, potato virus X, potatovirus Y, tobacco etch virus, tobacco rattle virus and tobacco mosaicvirus. Id.

[0137] (K) An insect-specific antibody or an immunotoxin derivedtherefrom. Thus, an antibody targeted to a critical metabolic functionin the insect gut would inactivate an affected enzyme, killing theinsect. Cf. Taylor et al., Abstract #497, SEVENTH INT'L SYMPOSIUM ONMOLECULAR PLANT-MICROBE INTERACTIONS (Edinburgh, Scotland, 1994)(enzymatic inactivation in transgenic tobacco via production ofsingle-chain antibody fragments).

[0138] (L) A virus-specific antibody. See, for example, Tavladoraki etal., Nature 366: 469 (1993), who show that transgenic plants expressingrecombinant antibody genes are protected from virus attack.

[0139] (M) A developmental-arrestive protein produced in nature by apathogen or a parasite. Thus, fungal endo alpha-1,4-D-polygalacturonasesfacilitate fungal colonization and plant nutrient release bysolubilizing plant cell wall homo-alpha-1,4-D-galacturonase. See Lamb etal., Bio/Technology 10: 1436 (1992). The cloning and characterization ofa gene which encodes a bean endopolygalacturonase-inhibiting protein isdescribed by Toubart et al., Plant J. 2: 367 (1992).

[0140] (N) A developmental-arrestive protein produced in nature by aplant. For example, Logemann et al., Bio/Technology 10: 305 (1992), haveshown that transgenic plants expressing the barley ribosome-inactivatinggene have an increased resistance to fungal disease.

[0141] (O) Genes involved in the Systemic Acquired Resistance (SAR)Response and/or the pathogenesis related genes. Briggs, S., CurrentBiology, 5(2) (1995).

[0142] (P) Antifungal genes (Cornelissen and Melchers, Pl. Physiol.101:709-712, (1993) and Parijs et al., Planta 183:258-264, (1991) andBushnell et al., Can. J. of Plant Path. 20(2):137-149 (1998).

[0143] (Q) Detoxification genes, such as for fumonisin, beauvericin,moniliformin and zearalenone and their structurally related derivatives.For example, see U.S. Pat. No. 5,792,931.

[0144] (R) Cystatin and cysteine proteinase inhibitors.

[0145] (S) Defensin genes. See WO03000863.

[0146] (T) Genes conferring resistance to nematodes, and in particularsoybean cyst nematodes. See e.g. PCT Application WO96/30517; PCTApplication WO93/19181, WO 03/033651 and Urwin et. al., Planta204:472-479 (1998).

[0147] (U) Genes that confer resistance to Phytophthora Root Rot, suchas the Rps 1, Rps 1-a, Rps 1-b, Rps 1-c, Rps 1-d, Rps 1-e, Rps 1-k, Rps2, Rps 3-a, Rps 3-b, Rps 3-c, Rps 4, Rps 5, Rps 6, Rps 7 and other Rpsgenes. See, for example, Shoemaker et al, Phytophthora Root RotResistance Gene Mapping in Soybean, Plant Genome IV Conference, SanDiego, Calif. (1995).

[0148] (V) Genes that confer resistance to Brown Stem Rot, such asdescribed in U.S. Pat. No. 5,689,035 and incorporated by reference forthis purpose.

[0149] 2. Transgenes that Confer Resistance to a Herbicide, for Example:

[0150] (A) A herbicide that inhibits the growing point or meristem, suchas an imidazolinone or a sulfonylurea. Exemplary genes in this categorycode for mutant ALS and AHAS enzyme as described, for example, by Lee etal., EMBO J. 7: 1241 (1988), and Miki et al., Theor. Appl. Genet. 80:449 (1990), respectively. See also, U.S. Pat. Nos. 5,605,011; 5,013,659;5,141,870; 5,767,361; 5,731,180; 5,304,732; 4,761,373; 5,331,107;5,928,937; and 5,378,824; and international publication WO 96/33270,which are incorporated herein by reference for this purpose.

[0151] (B) Glyphosate (resistance imparted by mutant5-enolpyruvl-3-phosphikimate synthase (EPSP) and aroA genes,respectively) and other phosphono compounds such as glufosinate(phosphinothricin acetyl transferase (PAT) and Streptomyceshygroscopicus phosphinothricin acetyl transferase (bar) genes), andpyridinoxy or phenoxy proprionic acids and cycloshexones (ACCaseinhibitor-encoding genes). See, for example, U.S. Pat. No. 4,940,835 toShah et al., which discloses the nucleotide sequence of a form of EPSPSwhich can confer glyphosate resistance. U.S. Pat. No. 5,627,061 to Barryet al. also describes genes encoding EPSPS enzymes. See also U.S. Pat.Nos. 6,248,876 B1; 6,040,497; 5,804,425; 5,633,435; 5,145,783;4,971,908; 5,312,910; 5,188,642; 4,940,835; 5,866,775; 6,225,114 B1;6,130,366; 5,310,667; 4,535,060; 4,769,061; 5,633,448; 5,510,471; Re.36,449; RE 37,287 E; and 5,491,288; and international publications WO97/04103; WO 97/04114; WO 00/66746; WO 01/66704; WO 00/66747 and WO00/66748, which are incorporated herein by reference for this purpose.Glyphosate resistance is also imparted to plants that express a genethat encodes a glyphosate oxido-reductase enzyme as described more fullyin U.S. Pat. Nos. 5,776,760 and 5,463,175, which are incorporated hereinby reference for this purpose. In addition glyphosate resistance can beimparted to plants by the over expression of genes encoding glyphosateN-acetyltransferase. See, for example, U.S. Application Serial Nos.60/244,385; 60/377,175 and 60/377,719. A DNA molecule encoding a mutantaroA gene can be obtained under ATCC accession No. 39256, and thenucleotide sequence of the mutant gene is disclosed in U.S. Pat. No.4,769,061 to Comai. European patent application No. 0 333 033 to Kumadaet al. and U.S. Pat. No. 4,975,374 to Goodman et al. disclose nucleotidesequences of glutamine synthetase genes which confer resistance toherbicides such as L-phosphinothricin. The nucleotide sequence of aphosphinothricin-acetyl-transferase gene is provided in European PatentNo. 0 242 246 and 0 242 236 to Leemans et al. De Greef et al.,Bio/Technology 7: 61 (1989), describe the production of transgenicplants that express chimeric bar genes coding for phosphinothricinacetyl transferase activity. See also, U.S. Pat. Nos. 5,969,213;5,489,520; 5,550,318; 5,874,265; 5,919,675; 5,561,236; 5,648,477;5,646,024; 6,177,616 B1; and 5,879,903, which are incorporated herein byreference for this purpose. Exemplary genes conferring resistance tophenoxy proprionic acids and cycloshexones, such as sethoxydim andhaloxyfop, are the Acc1-S1, Acc1-S2 and Acc1-S3 genes described byMarshall et al., Theor. Appl. Genet. 83: 435 (1992).

[0152] (C) A herbicide that inhibits photosynthesis, such as a triazine(psbA and gs+ genes) and a benzonitrile (nitrilase gene). Przibilla etal., Plant Cell 3: 169 (1991), describe the transformation ofChlamydomonas with plasmids encoding mutant psbA genes. Nucleotidesequences for nitrilase genes are disclosed in U.S. Pat. No. 4,810,648to Stalker, and DNA molecules containing these genes are available underATCC Accession Nos. 53435, 67441 and 67442. Cloning and expression ofDNA coding for a glutathione S-transferase is described by Hayes et al.,Biochem. J. 285:173 (1992).

[0153] (D) Acetohydroxy acid synthase, which has been found to makeplants that express this enzyme resistant to multiple types ofherbicides, has been introduced into a variety of plants (see, e.g.,Hattori et al. (1995) Mol Gen Genet 246:419). Other genes that confertolerance to herbicides include: a gene encoding a chimeric protein ofrat cytochrome P4507A1 and yeast NADPH-cytochrome P450 oxidoreductase(Shiota et al. (1994) Plant Physiol Plant Physiol 106:17), genes forglutathione reductase and superoxide dismutase (Aono et al. (1995) PlantCell Physiol 36:1687, and genes for various phosphotransferases (Dattaet al. (1992) Plant Mol Biol 20:619).

[0154] (E) Protoporphyrinogen oxidase (protox) is necessary for theproduction of chlorophyll, which is necessary for all plant survival.The protox enzyme serves as the target for a variety of herbicidalcompounds. These herbicides also inhibit growth of all the differentspecies of plants present, causing their total destruction. Thedevelopment of plants containing altered protox activity which areresistant to these herbicides are described in U.S. Pat. Nos. 6,288,306B1; 6,282,837 B1; and 5,767,373; and international publication WO01/12825, which are incorporated herein by reference for this purpose.

[0155] 3. Transgenes that Confer or Contribute to a Grain Trait, suchas:

[0156] (A) Modified fatty acid metabolism, for example, by

[0157] (1) Transforming a plant with an antisense gene of stearoyl-ACPdesaturase to increase stearic acid content of the plant. See Knultzonet al., Proc. Natl. Acad. Sci. USA 89: 2624 (1992),

[0158] (2) Elevating oleic acid via FAD-2 gene modification and/ordecreasing linolenic acid via FAD-3 gene modification (see U.S. Pat.Nos. 6,063,947; 6,323,392; and WO 93/11245),

[0159] (3) Altering conjugated linolenic or linoleic acid content, suchas in WO 01/12800,

[0160] (4) Modifying LEC1, AGP, Dek1, Superal1, and/or thioredoxin. Forexample, see WO 02/42424, WO 98/22604, WO 03/011015, U.S. Pat. No.6,423,886 and Rivera-Madrid, R. et. al. Proc. Natl. Acad. Sci.92:5620-5624 (1995).

[0161] (B) Decreased phytate content, for example, by the

[0162] (1) Introduction of a phytase-encoding gene would enhancebreakdown of phytate, adding more free phosphate to the transformedplant. For example, see Van Hartingsveldt et al., Gene 127: 87 (1993),for a disclosure of the nucleotide sequence of an Aspergillus nigerphytase gene.

[0163] (2) Introduction of a gene that reduces phytate content. Inmaize, this, for example, could be accomplished, by cloning and thenre-introducing DNA associated with one or more of the alleles, such asthe LPA alleles, identified in maize mutants characterized by low levelsof phytic acid, such as in Raboy et al., Maydica 35: 383 (1990) and/orby altering inositol kinase activity as in WO 02/059324, US2003/0009011,WO 03/027243, US2003/0079247 and WO 99/05298.

[0164] (C) Modified carbohydrate composition effected, for example, bytransforming plants with a gene coding for an enzyme that alters thebranching pattern of starch. See Shiroza et al., J. Bacteriol. 170: 810(1988) (nucleotide sequence of Streptococcus mutans fructosyltransferasegene), Steinmetz et al., Mol. Gen. Genet. 200: 220 (1985) (nucleotidesequence of Bacillus subtilis levansucrase gene), Pen et al.,Bio/Technology 10: 292 (1992) (production of transgenic plants thatexpress Bacillus licheniformis alpha-amylase), Elliot et al., PlantMolec. Biol. 21: 515 (1993) (nucleotide sequences of tomato invertasegenes), Søgaard et al., J. Biol. Chem. 268: 22480 (1993) (site-directedmutagenesis of barley alpha-amylase gene), and Fisher et al., PlantPhysiol. 102: 1045 (1993) (maize endosperm starch branching enzyme II).The fatty acid modification genes mentioned above may also be used toeffect starch content and/or composition through the interrelationshipof the starch and oil pathways.

[0165] (D) Altered antioxidant content or composition, such asalteration of tocopherol or tocotrienols. For example, see WO 00/68393involving the manipulation of antioxidant levels through alteration of aphytl prenyl transferase and WO 03/082899 through alteration of ahomogentisate geranyl geranyl transferase.

[0166] (E) Improved digestibility and/or starch extraction throughmodification of UDP-D-xylose 4-epimerase, Fragile 1 and 2, Ref1, HCHL,C4H, such as in WO 99/10498.

[0167] 4. Genes that Control Male-Sterility

[0168] (A) Introduction of a deacetylase gene under the control of atapetum-specific promoter and with the application of the chemicalN-Ac-PPT (WO 01/29237).

[0169] (B) Introduction of various stamen-specific promoters (WO92/13956, WO 92/13957).

[0170] (C) Introduction of the barnase and the barstar gene (Paul et al.Plant Mol. Biol. 19:611-622, 1992).

[0171] 5. Genes that create a site for site specific DNA integration.This includes the introduction of FRT sites that may be used in theFLP/FRT system and/or Lox sites that may be used in the Cre/Loxp system.For example, see Lyznik, et al., Site-Specific Recombination for GeneticEngineering in Plants, Plant Cell Rep (2003) 21:925-932 and WO 99/25821,which are hereby incorporated by reference. Other systems that may beused include the Gin recombinase of phage Mu (Maeser et al., 1991), thePin recombinase of E. coli (Enomoto et al., 1983), and the R/RS systemof the pSR1 plasmid (Araki et al., 1992).

[0172] 6. Genes that affect growth characteristics, such as droughttolerance and nitrogen utilization. For example, see WO 00/73475 wherewater use efficiency is modulated through alteration of malate.

[0173] Using XB04D4 to Develop Other Soybean Varieties

[0174] Soybean varieties such as XB04D4 are typically developed for usein seed and grain production. However, soybean varieties such as XB04D4also provide a source of breeding material that may be used to developnew soybean varieties. Plant breeding techniques known in the art andused in a soybean plant breeding program include, but are not limitedto, recurrent selection, mass selection, bulk selection, mass selection,backcrossing, pedigree breeding, open pollination breeding, restrictionfragment length polymorphism enhanced selection, genetic marker enhancedselection, making double haploids, and transformation. Oftencombinations of these techniques are used. The development of soybeanvarieties in a plant breeding program requires, in general, thedevelopment and evaluation of homozygous varieties. There are manyanalytical methods available to evaluate a new variety. The oldest andmost traditional method of analysis is the observation of phenotypictraits but genotypic analysis may also be used.

[0175] Using XB04D4 in a Breeding Program

[0176] This invention is directed to methods for producing a soybeanplant by crossing a first parent soybean plant with a second parentsoybean plant wherein either the first or second parent soybean plant isvariety XB04D4. The other parent may be any other soybean plant, such asa soybean plant that is part of a synthetic or natural population. Anysuch methods using soybean variety XB04D4 are part of this invention:selfing, sibbing, backcrosses, mass selection, pedigree breeding, bulkselection, hybrid production, crosses to populations, and the like.These methods are well known in the art and some of the more commonlyused breeding methods are described below. Descriptions of breedingmethods can be found in one of several reference books (e.g., Allard,Principles of Plant Breeding, 1960; Simmonds, Principles of CropImprovement, 1979; Sneep et al., 1979; Fehr, “Breeding Methods forCultivar Development”, Chapter 7, Soybean Improvement, Production andUses, 2^(nd) ed., Wilcox editor, 1987).

[0177] Pedigree Breeding

[0178] Pedigree breeding starts with the crossing of two genotypes, suchas XB04D4 and another soybean variety having one or more desirablecharacteristics that is lacking or which complements XB04D4. If the twooriginal parents do not provide all the desired characteristics, othersources can be included in the breeding population. In the pedigreemethod, superior plants are selfed and selected in successive filialgenerations. In the succeeding filial generations the heterozygouscondition gives way to homogeneous varieties as a result ofself-pollination and selection. Typically in the pedigree method ofbreeding, five or more successive filial generations of selfing andselection is practiced: F1→F2; F2→F3; F3→F4; F4→F₅, etc. After asufficient amount of inbreeding, successive filial generations willserve to increase seed of the developed variety. Preferably, thedeveloped variety comprises homozygous alleles at about 95% or more ofits loci.

[0179] In addition to being used to create a backcross conversion,backcrossing can also be used in combination with pedigree breeding. Asdiscussed previously, backcrossing can be used to transfer one or morespecifically desirable traits from one variety, the donor parent, to adeveloped variety called the recurrent parent, which has overall goodagronomic characteristics yet lacks that desirable trait or traits.However, the same procedure can be used to move the progeny toward thegenotype of the recurrent parent but at the same time retain manycomponents of the non-recurrent parent by stopping the backcrossing atan early stage and proceeding with selfing and selection. For example, asoybean variety may be crossed with another variety to produce a firstgeneration progeny plant. The first generation progeny plant may then bebackcrossed to one of its parent varieties to create a BC1 or BC2.Progeny are selfed and selected so that the newly developed variety hasmany of the attributes of the recurrent parent and yet several of thedesired attributes of the non-recurrent parent. This approach leveragesthe value and strengths of the recurrent parent for use in new soybeanvarieties.

[0180] Therefore, an embodiment of this invention is a method of makinga backcross conversion of soybean variety XB04D4, comprising the stepsof crossing a plant of soybean variety XB04D4 with a donor plantcomprising a desired trait, selecting an F1 progeny plant comprising thedesired trait, and backcrossing the selected F1 progeny plant to a plantof soybean variety XB04D4. This method may further comprise the step ofobtaining a molecular marker profile of soybean variety XB04D4 and usingthe molecular marker profile to select for a progeny plant with thedesired trait and the molecular marker profile of XB04D4. In oneembodiment the desired trait is a mutant gene or transgene present inthe donor parent.

[0181] Recurrent Selection and Mass Selection

[0182] Recurrent selection is a method used in a plant breeding programto improve a population of plants. XB04D4 is suitable for use in arecurrent selection program. The method entails individual plants crosspollinating with each other to form progeny. The progeny are grown andthe superior progeny selected by any number of selection methods, whichinclude individual plant, half-sib progeny, full-sib progeny and selfedprogeny. The selected progeny are cross pollinated with each other toform progeny for another population. This population is planted andagain superior plants are selected to cross pollinate with each other.Recurrent selection is a cyclical process and therefore can be repeatedas many times as desired. The objective of recurrent selection is toimprove the traits of a population. The improved population can then beused as a source of breeding material to obtain new varieties forcommercial or breeding use, including the production of a syntheticcultivar. A synthetic cultivar is the resultant progeny formed by theintercrossing of several selected varieties.

[0183] Mass selection is a useful technique when used in conjunctionwith molecular marker enhanced selection. In mass selection seeds fromindividuals are selected based on phenotype or genotype. These selectedseeds are then bulked and used to grow the next generation. Bulkselection requires growing a population of plants in a bulk plot,allowing the plants to self-pollinate, harvesting the seed in bulk andthen using a sample of the seed harvested in bulk to plant the nextgeneration. Also, instead of self pollination, directed pollinationcould be used as part of the breeding program.

[0184] Mutation Breeding

[0185] Mutation breeding is another method of introducing new traitsinto soybean variety XB04D4. Mutations that occur spontaneously or areartificially induced can be useful sources of variability for a plantbreeder. The goal of artificial mutagenesis is to increase the rate ofmutation for a desired characteristic. Mutation rates can be increasedby many different means including temperature, long-term seed storage,tissue culture conditions, radiation; such as X-rays, Gamma rays (e.g.cobalt 60 or cesium 137), neutrons, (product of nuclear fission byuranium 235 in an atomic reactor), Beta radiation (emitted fromradioisotopes such as phosphorus 32 or carbon 14), or ultravioletradiation (preferably from 2500 to 2900 nm), or chemical mutagens (suchas base analogues (5-bromo-uracil), related compounds (8-ethoxycaffeine), antibiotics (streptonigrin), alkylating agents (sulfurmustards, nitrogen mustards, epoxides, ethylenamines, sulfates,sulfonates, sulfones, lactones), azide, hydroxylamine, nitrous acid, oracridines. Once a desired trait is observed through mutagenesis thetrait may then be incorporated into existing germplasm by traditionalbreeding techniques. Details of mutation breeding can be found in“Principals of Cultivar Development” Fehr, 1993 Macmillan PublishingCompany the disclosure of which is incorporated herein by reference. Inaddition, mutations created in other soybean plants may be used toproduce a backcross conversion of XB04D4 that comprises such mutation.

[0186] Breeding with Molecular Markers

[0187] Molecular markers, which includes markers identified through theuse of techniques such as Isozyme Electrophoresis, Restriction FragmentLength Polymorphisms (RFLPs), Randomly Amplified Polymorphic DNAs(RAPDs), Arbitrarily Primed Polymerase Chain Reaction (AP-PCR), DNAAmplification Fingerprinting (DAF), Sequence Characterized AmplifiedRegions (SCARs), Amplified Fragment Length Polymorphisms (AFLPs), SimpleSequence Repeats (SSRs) and Single Nucleotide Polymorphisms (SNPs), maybe used in plant breeding methods utilizing XB04D4.

[0188] Isozyme Electrophoresis and RFLPs have been widely used todetermine genetic composition. Shoemaker and Olsen, ((1993) MolecularLinkage Map of Soybean (Glycine max L. Merr.). p. 6.131-6.138. In S. J.O'Brien (ed.) Genetic Maps: Locus Maps of Complex Genomes. Cold SpringHarbor Laboratory Press. Cold Spring Harbor, N.Y.), developed amolecular genetic linkage map that consisted of 25 linkage groups withabout 365 RFLP, 11 RAPD (random amplified polymorphic DNA), threeclassical markers, and four isozyme loci. See also, Shoemaker R. C. 1994RFLP Map of Soybean. P. 299-309 In R. L. Phillips and I. K. Vasil (ed.)DNA-based markers in plants. Kluwer Academic Press Dordrecht, theNetherlands.

[0189] SSR technology is currently the most efficient and practicalmarker technology; more marker loci can be routinely used and morealleles per marker locus can be found using SSRs in comparison to RFLPs.For example Diwan and Cregan, described a highly polymorphicmicrosatellite loci in soybean with as many as 26 alleles. (Diwan, N.,and P. B. Cregan 1997 Automated sizing of fluorescent-labeled simplesequence repeat (SSR) markers to assay genetic variation in SoybeanTheor. Appl. Genet. 95:220-225.) Single Nucleotide Polymorphisms mayalso be used to identify the unique genetic composition of the inventionand progeny varieties retaining that unique genetic composition. Variousmolecular marker techniques may be used in combination to enhanceoverall resolution.

[0190] Soybean DNA molecular marker linkage maps have been rapidlyconstructed and widely implemented in genetic studies. One such study isdescribed in Cregan et. al, “An Integrated Genetic Linkage Map of theSoybean Genome” Crop Science 39:1464-1490 (1999). Sequences and PCRconditions of SSR Loci in Soybean as well as the most current geneticmap may be found in Soybase on the world wide web.

[0191] One use of molecular markers is Quantitative Trait Loci (QTL)mapping. QTL mapping is the use of markers, which are known to beclosely linked to alleles that have measurable effects on a quantitativetrait. Selection in the breeding process is based upon the accumulationof markers linked to the positive effecting alleles and/or theelimination of the markers linked to the negative effecting alleles fromthe plant's genome.

[0192] Molecular markers can also be used during the breeding processfor the selection of qualitative traits. For example, markers closelylinked to alleles or markers containing sequences within the actualalleles of interest can be used to select plants that contain thealleles of interest during a backcrossing breeding program. The markerscan also be used to select for the genome of the recurrent parent andagainst the genome of the donor parent. Using this procedure canminimize the amount of genome from the donor parent that remains in theselected plants. It can also be used to reduce the number of crossesback to the recurrent parent needed in a backcrossing program. The useof molecular markers in the selection process is often called geneticmarker enhanced selection.

[0193] Production of Double Haploids

[0194] The production of double haploids can also be used for thedevelopment of plants with a homozygous phenotype in the breedingprogram. For example, a soybean plant for which XB04D4 is a parent canbe used to produce double haploid plants. Double haploids are producedby the doubling of a set of chromosomes (1N) from a heterozygous plantto produce a completely homozygous individual. For example, see Wan etal., “Efficient Production of Doubled Haploid Plants Through ColchicineTreatment of Anther-Derived Maize Callus”, Theoretical and AppliedGenetics, 77:889-892, 1989 and US2003/0005479. This can be advantageousbecause the process omits the generations of selfing needed to obtain ahomozygous plant from a heterozygous source.

[0195] Thus, an embodiment of this invention is a process for making asubstantially homozygous XB04D04 progeny plant by producing or obtaininga seed from the cross of XB04D4 and another soybean plant and applyingdouble haploid methods to the F1 seed or F1 plant or to any successivefilial generation. Based on studies in maize and currently beingconducted in soybean, such methods would decrease the number ofgenerations required to produce a variety with similar genetics orcharacteristics to XB04D4. See Bernardo, R. and Kahler, A. L., Theor.Appl. Genet. 102:986-992, 2001.

[0196] In particular, a process of making seed retaining the molecularmarker profile of soybean variety XB04D4 is contemplated, such processcomprising obtaining or producing F1 seed for which soybean varietyXB04D4 is a parent, inducing doubled haploids to create progeny withoutthe occurrence of meiotic segregation, obtaining the molecular markerprofile of soybean variety XB04D4, and selecting progeny that retain themolecular marker profile of XB04D4.

[0197] Use of XB04D4 in Tissue Culture

[0198] This invention is also directed to the use of variety XB04D4 intissue culture. Tissue culture of various tissues of soybeans andregeneration of plants therefrom is well known and widely published. Forexample, reference may be had to Komatsuda, T. et al., “Genotype XSucrose Interactions for Somatic Embryogenesis in Soybean,” Crop Sci.31:333-337 (1991); Stephens, P. A. et al., “Agronomic Evaluation ofTissue-Culture-Derived Soybean Plants,” Theor. Appl. Genet. (1991)82:633-635; Komatsuda, T. et al., “Maturation and Germination of SomaticEmbryos as Affected by Sucrose and Plant Growth Regulators in SoybeansGlycine gracilis Skvortz and Glycine max (L.) Merr.,” Plant Cell, Tissueand Organ Culture, 28:103-113 (1992); Dhir, S. et al., “Regeneration ofFertile Plants from Protoplasts of Soybean (Glycine max L. Merr.):Genotypic Differences in Culture Response,” Plant Cell Reports (1992)11:285-289; Pandey, P. et al., “Plant Regeneration from Leaf andHypocotyl Explants of Glycine wightii (W. and A.) VERDC. var.longicauda,” Japan J. Breed. 42:1-5 (1992); and Shetty, K., et al.,“Stimulation of In Vitro Shoot Organogenesis in Glycine max (Merrill.)by Allantoin and Amides,” Plant Science 81:(1992) 245-251; as well asU.S. Pat. No. 5,024,944, issued Jun. 18, 1991 to Collins et al. and U.S.Pat. No. 5,008,200, issued Apr. 16, 1991 to Ranch et al., thedisclosures of which are hereby incorporated herein in their entirety byreference. Thus, another aspect of this invention is to provide cellswhich upon growth and differentiation produce soybean plants having thephysiological and morphological characteristics of soybean varietyXB04D4.

[0199] Progeny Plants

[0200] All plants produced by the use of the methods described hereinand that retain the unique genetic or trait combinations of XB04D4 arewithin the scope of the invention. Progeny of the breeding methodsdescribed herein may be characterized in any number of ways, such as bytraits retained in the progeny, pedigree and/or molecular markers.Combinations of these methods of characterization may be used.

[0201] Breeders of ordinary skill in the art have developed the conceptof an “essentially derived variety”, which is defined in 7 U.S.C. §2104(a)(3) of the Plant Variety Protection Act and is herebyincorporated by reference. Varieties and plants that are essentiallyderived from XB04D4 are within the scope of the invention.

[0202] Pedigree is a method used by breeders of ordinary skill in theart to describe the varieties. Varieties that are more closely relatedby pedigree are likely to share common genotypes and combinations ofphenotypic characteristics. All breeders of ordinary skill in the artmaintain pedigree records of their breeding programs. These pedigreerecords contain a detailed description of the breeding process,including a listing of all parental varieties used in the breedingprocess and information on how such variety was used. One embodiment ofthis invention is progeny plants and parts thereof with at least oneancestor that is XB04D4, and more specifically, where the pedigree ofthe progeny includes 1, 2, 3, 4, and/or 5 or less breeding crosses to asoybean plant other than XB04D04 or a plant that has XB04D4 as a parentor other progenitor. A breeder of ordinary skill in the art would knowif XB04D4 were used in the development of a progeny variety, and wouldalso know how many crosses to a variety other than XB04D4 or varietywith XB04D4 as a parent or other progenitor were made in the developmentof any progeny variety.

[0203] Molecular markers also provide a means by which those of ordinaryskill in the art characterize the similarity or differences of twovarieties. Using the breeding methods described herein, one can developindividual plants, plant cells, and populations of plants that retain atleast 25%, 30%, 35%, 40%, 45%, 50%, 55%, 60%, 61%, 62%, 63%, 64%, 65%,66%, 67%, 68%, 69%, 70%, 71%, 72%, 73%, 74%, 75%, 76%, 77%, 78%, 79%,80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%,94%, 95%, 96%, 97%, 98%, 99% or 99.5% genetic contribution from soybeanvariety XB04D04, as measured by either percent identity or percentsimilarity. In pedigree analysis the percentage genetic contribution maynot be actually known, but on average 50% of the starting germplasmwould be expected to be passed to the progeny variety after one cross toanother variety, 25% after another cross to a different variety, and soon. With backcrossing, the expected contribution of XB04D4 after 2, 3, 4and 5 doses (or 1, 2, 3 and 4 backcrosses) would be 75%, 87.5%, 93.75%and 96.875% respectively. Actual genetic contribution may be much higherthan the genetic contribution expected by pedigree, especially ifmolecular markers are used in selection. Molecular markers could also beused to confirm and/or determine the pedigree of the progeny variety.

[0204] Traits are also used by those of ordinary skill in the art tocharacterize progeny. Traits are commonly evaluated at a significancelevel, such as a 1%, 5% or 10% significance level, when measured inplants grown in the same environmental conditions. For example, abackcross conversion of XB04D04 may be characterized as having the samemorphological and physiological traits as XB04D4. The traits used forcomparison may be any or all of the traits shown in Table 1 or Table 2.Environmental conditions should be appropriate for the traits or traitsbeing evaluated. Sufficient selection pressure should be present foroptimum measurement of traits of interest such as herbicide, insect ordisease resistance. Similarly, an introgressed trait conversion ofXB04D4 for resistance, such as herbicide resistance, should not becompared to XB04D4 in the presence of the herbicide when comparingnon-resistance related traits such as plant height and yield.

[0205] The population of plants produced at each and any cycle ofbreeding is also an embodiment of the invention, and on average eachsuch population would predictably consist of plants containingapproximately 50% of its genes from variety XB04D4 in the first breedingcycle, 25% of its genes from variety XB04D4 in the second breedingcycle, 12.5% of its genes from variety XB04D4 in the third breedingcycle, 6.25% in the fourth breeding cycle, and so on. However, in eachcase the use of variety XB04D4 provides a benefit, because linkagegroups of XB04D4 are retained in the progeny varieties. Specifically, anembodiment of the invention is a process for making a population ofXB04D4 progeny plants comprising obtaining or producing a firstgeneration progeny seed comprising the plant of XB04D4 as a parent,growing said first generation progeny seed to produce first generationplants, obtaining self or sib pollinated seed from said first generationplants, and growing the self or sib pollinated seed to obtain apopulation of XB04D4 progeny plants.

[0206] The population of XB04D4 progeny soybean plants produced by thismethod will retain the expected genetic contribution of XB04D4 describedabove. A variety selected from the population of XB04D4 progeny plantsproduced by this method is an embodiment, and such variety may befurther characterized by its molecular marker identity or similarity toXB04D4.

[0207] In this manner, the invention encompasses a process for making asubstantially homozygous XB04D4 progeny plant comprising the steps ofobtaining or producing a first generation progeny seed wherein a parentof said first generation progeny seed is a plant of variety XB04D4,growing said first generation progeny seed to produce a first generationplant and obtaining self or sib pollinated seed from said firstgeneration soybean plant, and producing successive filial generations toobtain a substantially homozygous XB04D4 progeny plant. Also anembodiment of this invention is the substantially homozygous XB04D4progeny plant produced by this method.

Deposits

[0208] Applicant(s) will make a deposit of at least 2500 seeds ofSoybean Variety XB04D4 with the American Type Culture Collection (ATCC),Manassas, Va. 20110 USA, ATCC Deposit No. ______. The seeds depositedwith the ATCC on ______ will be taken from the deposit maintained byPioneer Hi-Bred International, Inc., 800 Capital Square, 400 LocustStreet, Des Moines, Iowa 50309-2340 since prior to the filing date ofthis application. Access to this deposit will be available during thependency of the application to the Commissioner of Patents andTrademarks and persons determined by the Commissioner to be entitledthereto upon request. Upon allowance of any claims in the application,the Applicant(s) will make the deposit available to the public pursuantto 37 CFR 1.808. This deposit of Soybean Variety XB04D4 will bemaintained in the ATCC depository, which is a public depository, for aperiod of 30 years, or 5 years after the most recent request, or for theenforceable life of the patent, whichever is longer, and will bereplaced if it becomes nonviable during that period. Additionally,Applicant(s) have or will satisfy all the requirements of 37 C.F.R.§§1.801-1.809, including providing an indication of the viability of thesample upon deposit. Applicant(s) have no authority to waive anyrestrictions imposed by law on the transfer of biological material orits transportation in commerce. Applicant(s) do not waive anyinfringement of their rights granted under this patent or under thePlant Variety Protection Act (7 USC 2321 et seq.).

[0209] All publications, patents and patent applications mentioned inthe specification are indicative of the level of those skilled in theart to which this invention pertains. All such publications, patents andpatent applications are incorporated by reference herein for the purposecited to the same extent as if each was specifically and individuallyindicated to be incorporated by reference herein.

[0210] The foregoing invention has been described in detail by way ofillustration and example for purposes of clarity and understanding. Asis readily apparent to one skilled in the art, the foregoing are onlysome of the methods and compositions that illustrate the embodiments ofthe foregoing invention. It will be apparent to those of ordinary skillin the art that variations, changes, modifications and alterations maybe applied to the compositions and/or methods described herein withoutdeparting from the true spirit, concept and scope of the invention.

What is claimed is:
 1. Seed of soybean variety XB04D4, representativeseed of said soybean variety XB04D4 having been deposited under ATCCAccession No. ______.
 2. A soybean plant, or a part thereof, produced bygrowing the seed of claim
 1. 3. The soybean plant part of claim 2wherein said part is pollen.
 4. The soybean plant part of claim 2wherein said part is an ovule.
 5. A tissue culture of protoplasts orregenerable cells from the plant of claim
 2. 6. A tissue cultureaccording to claim 5, wherein cells or protoplasts of the tissue cultureare obtained from plant tissue selected from the group consisting of:leaf, pollen, cotyledon, hypocotyl, embryos, root, pod, flower, shootand stem.
 7. A soybean plant regenerated from the tissue culture ofclaim 5 and having all the morphological and physiologicalcharacteristics of soybean variety XB04D4, representative seed of saidsoybean variety XB04D4 having been deposited under ATCC Accession No.______.
 8. A method for producing a progeny soybean plant comprising:crossing the plant of claim 2 with a different soybean plant, harvestingthe resultant soybean seed, and growing a soybean plant.